Class I molecules of the major histocompatibility complex class (MHC) consist of a highly polymorphic heavy chain complexed to B2 microglobulin. Class I molecules are expressed on virtually all cell types. Their sole function is to bind antigens and present them to T cells bearing CD8 molecules. These T cells are known as cytotoxic T lymphocytes (CTLs) due to their ability to lyse histocompatible cells in an antigen specific manner. CTLs play a critical role in eradicating intracellular pathogens and tumors. On the negative side, they are involved in organ rejection, and in many autoimmune dyscrasias. There has been rapid progress in understanding the physical nature of the antigen-class I complex and in how antigens become associated with class I molecules in cells. Based in part on results from this laboratory, it is now apparent that antigens present in the cytosol are translocated into the exocytic compartment where they bind class I molecules which carry them to the cell surface for CTL recognition. In the past year we have continued our studies on antigen processing, which can be defined as the structural modification and trafficking of protein antigens that enable the determinants recognized by CTLs (often buried in native proteins) to interact with MHC molecules in the proper subcellular compartment. We have focused on the following questions. In which exocytic compartment does association with class I molecules occur? Are there signals for targeting proteins into the cytosolic antigen processing pathway? Are there cellular proteins which function to facilitate the association of class I molecules with antigen? Can extracellular proteins be targeted to the cytosolic antigen processing pathway?